Summary of Work: The glucose analogue 2-fluoro-2-deoxyglucose (FDG) has been successfully utilized to image, stage, and evaluate tumor progression. While it is generally believed that FDG uptake is directly correlated with tumor glucose metabolism, the precise molecular mechanisms behind this process are not fully understood. Several human tumors have been shown to express the glucose transporter isoform GLUT- 1. To evaluate the relationship between GLUT-1 expression and FDG uptake, we have studied three human cancer cell lines derived from breast, liver, and an epidermoid carcinoma (T47D, HepG2 and A431, respectively) and correlated the results with the amount of GLUT-1 mRNA, and protein expression. We have previously shown that when these cells are grown as subcutaneous xenografts in nude mice, T47D cells have the highest [3H]DG uptake but the lowest amount of GLUT-1 mRNA. To evaluate FDG uptake in the absence of confounding variables (such as tumor blood flow or necrosis), we have now studied cells grown in culture. Cells were studied in different phases of growth. When cells were growing exponentially, FDG uptake in T47D was 50% higher than in A431 and 100% higher than in HepG2. When cells were confluent, T47D cells still showed higher uptake, but all three cell lines showed a reduction of approximately 50% in the rate of FDG uptake. Northern blot analysis of RNA extracted from exponentially growing cells showed that Beta-actin- normalized levels for GLUT-I in T47D cells were approximately 90% lower than in A431 and HepG2 cells. None of the other facilitative glucose transporter isoforms (GLUT 2-5) mRNAs were present at detectable levels in any of the cells. Western blot analysis showed a very small amount of immunodetectable GLUT-1 protein in T47D compared to A431 and HepG2 (66% and 83% lower, respectively). Our results show that in this system, where direct determinations of FDG uptake by cancer cells are made, the amount of GLUT-1 present on the cell membranes is not correlated with the level of FDG uptake.